UDP‐N‐Acetylglucosamine:Dolichyl‐Phosphate N‐Acetylglucosamine‐1‐Phosphate Transferase is Amplified in Tunicamycin‐Resistant Soybean Cells

• Published in: European journal of biochemistry Vol. 233; no. 2; pp. 458 - 466
• Main Authors: Zeng, Yucheng, Elbein, Alan D.
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Science Ltd 15.10.1995
• Subjects: amplification of GlcNAclP transferase; Anti-Bacterial Agents - pharmacology; Cell Line; complex carbohydrates; Drug Resistance, Microbial; Glycine max; Glycosylation; glycosylation‐altered soybean cells; N‐linked glycosylation; Transferases (Other Substituted Phosphate Groups) - metabolism; Tunicamycin - pharmacology; tunicamycin‐resistant
• ISSN: 0014-2956; 1432-1033
• DOI: 10.1111/j.1432-1033.1995.458_2.x

A tunicamycin‐resistant soybean cell line was developed by gradually increasing the concentration of tunicamycin in the growth medium. At the final stage, the resistant cells could survive in media containing 60 μg/ml of tunicamycin, whereas normal cells show a greatly retarded growth rate at 0.5 μg/ml of antibiotic. The tunicamycin‐resistant cells had a greater than 40‐fold increase in the activity of the enzyme UDP‐GlcNAc:dolichyl‐P GlcNAclP transferase, a 2–3‐fold increase in the activity of dolichyl‐P ‐mannose synthase, but no increase in the activities of other enzymes of the lipid‐linked saccharide pathway such as dolichyl‐P ‐glucose synthase or mannosyl transferases. There was also no change in the activities of the glycoprotein‐processing enzymes, glucosidase I or glucosidase II, as compared to wild‐type cells. The increase in GlcNAclP transferase was due to an increased production of enzyme, as seen by a dramatic increase in the amount of a 39–kDa protein, which is presumed to be this enzyme protein. The GlcNAclP transferase from tunicamycin‐resistant cells was equally sensitive to tunicamycin as was the wild‐type enzyme, but was considerably more labile to temperatures above 30°C. The activity in tunicamycin‐resistant cells was greatly stimulated by exogenous dolichyl‐P. The spectrum of oligosaccharides from labeled lipid‐linked oligosaccharides was similar in wild‐type and tunicamycin‐resistant soybean cells, but the resistant cells had significantly greater amounts of the shorter and much lower amounts of the larger‐sized oligosaccharides.